Nano silver treatment is effective in reducing bacterial contaminations ofAraucaria excelsaR. Br. var.glaucaexplants

Abstract

The downside of plant tissue culture techniques is an unwanted microbial contamination. Elimination of contaminants is the first step of any successful investigation on plant tissue culture. Preliminary experiments on Araucaria excelsa R. Br. var. glauca (Norfolk-Island pine) (syn.: A. heterophylla) showed that most common decontaminants could not successfully eliminate the contamination. Therefore, nano silver (NS) colloids were evaluated for controlling contamination. Treatments were included soaking the explants in NS solution or adding NS to the culture medium. Explants were cultured on MS medium supplemented with appropriate growth regulators for their establishment. Results showed that surface sterilization followed by treatment with 200 mg l-1 of NS with soaking time of 180 min reduced the bacterial contamination from 61.5% to 11.3% and adding 400 mg l-1 NS to the medium reduced the bacterial contamination from 81.25% to 18.75%. Nano silver could be applied without adverse effects on plant growth and development. This is the first report on in vitro establishment of A. excelsa R. Br. using NS to reduce bacterial infections.

RAPD Fingerprint to Appraise the Genetic Fidelity of In Vitro Propagated Araucaria excelsa R. Br. var. glauca Plantlets

“…glauca were chosen for this study. Pieces of orthotropic stem were cut from the 8 cm upper half and prewashed in tap water for 3 h. Owing to high contamination rate observed nano silver [19,20] was used to eradicate the internal bacteria with a concentration of 500 lm/ml under reduced pressure (300 mm Hg in 5 min).Then, explants were treated with 70% ethanol for 3 min and 15% Clorox (containing 5.25% sodium hypochlorite) with 0.2% household detergent for at least 10-20 min for surface sterilization, and then rinsed six times with sterilized distilled water. The stem pieces were then cut into 7-9 mm long explant and placed with their proximal ends on MS [21] basal medium with 3.0% sucrose and 0.8% agar.…”

In Vitro Rooting of Araucaria Excelsa R. Br. Var. Glauca Using Agrobacterium Rhizogenes

“…In most cases, application of 500 µg/ml nano silver for 5-10 min before surface sterilization reduced the contamination to less than 18% [16]. Apical parts of orthotropic stems can be used for good multiplication (Fig.…”

“…The potting medium consisted of a 1:1:1 (in volume) of loam, peat moss, and perlite and it was fertilized by Kristalon solution (1/1000, each 15 days). Four cm of apical stem segments of main stem were cut from seedlings and kept under running tap-water for 2 h. Because of high contamination rate, nano silver (NS) with concentration of 500 µg/ml under reduced pressure (300 mm Hg in 5 min) [16,17] was used to eradicate bacterial contamination. Then, the explants were treated with 70% ethanol for 3 min and 15% Clorox (containing 5.25% sodium hypochlorite) with 0.2% detergent for 10-20 min for surface sterilization, and then rinsed six times with sterilized distilled water.…”

“…[7]. Burrows et al (1988) made further efforts and used 2 years old seedlings and 20 years old coppices of A. cunninghamii as sources of explants and their investigation demonstrated that Murashige and Skoog's (MS) medium [16] supplemented with 0.1, 0.01 and 0.001 µM benzyl amino purine (BAP) had no significant effect on bud development, whereas concentrations more than 1 µM BAP caused abnormality of axillary buds. Furthermore, they found that the use of gibberellic acid (GA3) for elongation of shoots was not successful [8].…”

Micropropagation of Araucaria excelsa R. Br. var. glauca Carrière from orthotropic stem explants

“…Orthotropic stems of approximately 17 cm long were removed from the mother plants and after removing the apical bud, were divided into three parts (including upper, middle and lower segments of semihardwood orthotropic stems each 5 cm long) and were kept under running tap-water for 2 h. Because of high contamination, nano silver was used to eradicate the internal bacteria at concentration of 500 μg/ml under reduced pressure (300 mm Hg in 5 min) (Sarmast et al 2011). Then the explants were treated with 70 % ethanol for 3 min and 15 % Clorox (containing 5.25 % sodium hypochlorite) with 0.2 % detergent for at least 10-20 min for surface sterilization, and then rinsed six times with sterilized distilled water.…”